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- EMDB-27096: Structure of a Xenopus Nucleosome with Widom 601 DNA -

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Basic information

Entry
Database: EMDB / ID: EMD-27096
TitleStructure of a Xenopus Nucleosome with Widom 601 DNA
Map dataNucleosome
Sample
  • Complex: Nucleosome
    • Protein or peptide: Histone H3
    • Protein or peptide: Histone H4
    • Protein or peptide: Histone H2A
    • Protein or peptide: Histone H2B
    • DNA: Widom 601 DNA (146-MER)
    • DNA: Widom 601 DNA (146-MER)
Keywordsmeiosis / recombination / chromosome axis / nucleosome / PHD / winged helix / DNA BINDING PROTEIN-DNA complex
Biological speciesSaccharomyces cerevisiae (brewer's yeast) / Xenopus laevis (African clawed frog) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.58 Å
AuthorsGu Y / Ur SN / Milano CR / Tromer EC / Vale-Silva LA / Hochwagen A / Corbett KD
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM144121 United States
CitationJournal: EMBO J / Year: 2024
Title: Chromatin binding by HORMAD proteins regulates meiotic recombination initiation.
Authors: Carolyn R Milano / Sarah N Ur / Yajie Gu / Jessie Zhang / Rachal Allison / George Brown / Matthew J Neale / Eelco C Tromer / Kevin D Corbett / Andreas Hochwagen /
Abstract: The meiotic chromosome axis coordinates chromosome organization and interhomolog recombination in meiotic prophase and is essential for fertility. In S. cerevisiae, the HORMAD protein Hop1 mediates ...The meiotic chromosome axis coordinates chromosome organization and interhomolog recombination in meiotic prophase and is essential for fertility. In S. cerevisiae, the HORMAD protein Hop1 mediates the enrichment of axis proteins at nucleosome-rich islands through a central chromatin-binding region (CBR). Here, we use cryoelectron microscopy to show that the Hop1 CBR directly recognizes bent nucleosomal DNA through a composite interface in its PHD and winged helix-turn-helix domains. Targeted disruption of the Hop1 CBR-nucleosome interface causes a localized reduction of axis protein binding and meiotic DNA double-strand breaks (DSBs) in axis islands and leads to defects in chromosome synapsis. Synthetic effects with mutants of the Hop1 regulator Pch2 suggest that nucleosome binding delays a conformational switch in Hop1 from a DSB-promoting, Pch2-inaccessible state to a DSB-inactive, Pch2-accessible state to regulate the extent of meiotic DSB formation. Phylogenetic analyses of meiotic HORMADs reveal an ancient origin of the CBR, suggesting that the mechanisms we uncover are broadly conserved.
History
DepositionMay 24, 2022-
Header (metadata) releaseJun 7, 2023-
Map releaseJun 7, 2023-
UpdateApr 10, 2024-
Current statusApr 10, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_27096.png

Downloads & links

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Map

FileDownload / File: emd_27096.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationNucleosome
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.1 Å/pix.
x 256 pix.
= 281.6 Å		1.1 Å/pix.
x 256 pix.
= 281.6 Å		1.1 Å/pix.
x 256 pix.
= 281.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.1 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.22
Minimum - Maximum-0.41115722 - 1.2785027
Average (Standard dev.)-0.0008284051 (±0.03406025)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 281.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_27096_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_27096_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Nucleosome

EntireName: Nucleosome
Components
  • Complex: Nucleosome
    • Protein or peptide: Histone H3
    • Protein or peptide: Histone H4
    • Protein or peptide: Histone H2A
    • Protein or peptide: Histone H2B
    • DNA: Widom 601 DNA (146-MER)
    • DNA: Widom 601 DNA (146-MER)

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Supramolecule #1: Nucleosome

SupramoleculeName: Nucleosome / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast)
Molecular weightTheoretical: 200 KDa

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Macromolecule #1: Histone H3

MacromoleculeName: Histone H3 / type: protein_or_peptide / ID: 1 / Details: GenBank:CAD89679.1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Molecular weightTheoretical: 15.30393 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
ARTKQTARKS TGGKAPRKQL ATKAARKSAP ATGGVKKPHR YRPGTVALRE IRRYQKSTEL LIRKLPFQRL VREIAQDFKT DLRFQSSAV MALQEASEAY LVALFEDTNL CAIHAKRVTI MPKDIQLARR IRGERA

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Macromolecule #2: Histone H4

MacromoleculeName: Histone H4 / type: protein_or_peptide / ID: 2 / Details: GenBank:NP_001087926.1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Molecular weightTheoretical: 11.263231 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SGRGKGGKGL GKGGAKRHRK VLRDNIQGIT KPAIRRLARR GGVKRISGLI YEETRGVLKV FLENVIRDAV TYTEHAKRKT VTAMDVVYA LKRQGRTLYG FGG

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Macromolecule #3: Histone H2A

MacromoleculeName: Histone H2A / type: protein_or_peptide / ID: 3 / Details: GenBank:CAD89676.1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Molecular weightTheoretical: 13.978241 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
SGRGKQGGKT RAKAKTRSSR AGLQFPVGRV HRLLRKGNYA ERVGAGAPVY LAAVLEYLTA EILELAGNAA RDNKKTRIIP RHLQLAVRN DEELNKLLGR VTIAQGGVLP NIQSVLLPKK TESSKSAKSK

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Macromolecule #4: Histone H2B

MacromoleculeName: Histone H2B / type: protein_or_peptide / ID: 4 / Details: GenBank:CAD89678.1 / Number of copies: 2 / Enantiomer: LEVO
Source (natural)Organism: Xenopus laevis (African clawed frog)
Molecular weightTheoretical: 13.524752 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
AKSAPAPKKG SKKAVTKTQK KDGKKRRKTR KESYAIYVYK VLKQVHPDTG ISSKAMSIMN SFVNDVFERI AGEASRLAHY NKRSTITSR EIQTAVRLLL PGELAKHAVS EGTKAVTKYT SAK

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Macromolecule #5: Widom 601 DNA (146-MER)

MacromoleculeName: Widom 601 DNA (146-MER) / type: dna / ID: 5 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 45.27484 KDa
SequenceString: (DA)(DC)(DA)(DG)(DG)(DA)(DT)(DG)(DT)(DA) (DT)(DA)(DT)(DA)(DT)(DC)(DT)(DG)(DA)(DC) (DA)(DC)(DG)(DT)(DG)(DC)(DC)(DT)(DG) (DG)(DA)(DG)(DA)(DC)(DT)(DA)(DG)(DG)(DG) (DA) (DG)(DT)(DA)(DA)(DT)(DC) ...String:
(DA)(DC)(DA)(DG)(DG)(DA)(DT)(DG)(DT)(DA) (DT)(DA)(DT)(DA)(DT)(DC)(DT)(DG)(DA)(DC) (DA)(DC)(DG)(DT)(DG)(DC)(DC)(DT)(DG) (DG)(DA)(DG)(DA)(DC)(DT)(DA)(DG)(DG)(DG) (DA) (DG)(DT)(DA)(DA)(DT)(DC)(DC)(DC) (DC)(DT)(DT)(DG)(DG)(DC)(DG)(DG)(DT)(DT) (DA)(DA) (DA)(DA)(DC)(DG)(DC)(DG)(DG) (DG)(DG)(DG)(DA)(DC)(DA)(DG)(DC)(DG)(DC) (DG)(DT)(DA) (DC)(DG)(DT)(DG)(DC)(DG) (DT)(DT)(DT)(DA)(DA)(DG)(DC)(DG)(DG)(DT) (DG)(DC)(DT)(DA) (DG)(DA)(DG)(DC)(DT) (DG)(DT)(DC)(DT)(DA)(DC)(DG)(DA)(DC)(DC) (DA)(DA)(DT)(DT)(DG) (DA)(DG)(DC)(DG) (DG)(DC)(DC)(DT)(DC)(DG)(DG)(DC)(DA)(DC) (DC)(DG)(DG)(DG)(DA)(DT) (DT)(DC)(DT) (DC)(DC)(DA)

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Macromolecule #6: Widom 601 DNA (146-MER)

MacromoleculeName: Widom 601 DNA (146-MER) / type: dna / ID: 6 / Number of copies: 1 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 44.85657 KDa
SequenceString: (DT)(DG)(DG)(DA)(DG)(DA)(DA)(DT)(DC)(DC) (DC)(DG)(DG)(DT)(DG)(DC)(DC)(DG)(DA)(DG) (DG)(DC)(DC)(DG)(DC)(DT)(DC)(DA)(DA) (DT)(DT)(DG)(DG)(DT)(DC)(DG)(DT)(DA)(DG) (DA) (DC)(DA)(DG)(DC)(DT)(DC) ...String:
(DT)(DG)(DG)(DA)(DG)(DA)(DA)(DT)(DC)(DC) (DC)(DG)(DG)(DT)(DG)(DC)(DC)(DG)(DA)(DG) (DG)(DC)(DC)(DG)(DC)(DT)(DC)(DA)(DA) (DT)(DT)(DG)(DG)(DT)(DC)(DG)(DT)(DA)(DG) (DA) (DC)(DA)(DG)(DC)(DT)(DC)(DT)(DA) (DG)(DC)(DA)(DC)(DC)(DG)(DC)(DT)(DT)(DA) (DA)(DA) (DC)(DG)(DC)(DA)(DC)(DG)(DT) (DA)(DC)(DG)(DC)(DG)(DC)(DT)(DG)(DT)(DC) (DC)(DC)(DC) (DC)(DG)(DC)(DG)(DT)(DT) (DT)(DT)(DA)(DA)(DC)(DC)(DG)(DC)(DC)(DA) (DA)(DG)(DG)(DG) (DG)(DA)(DT)(DT)(DA) (DC)(DT)(DC)(DC)(DC)(DT)(DA)(DG)(DT)(DC) (DT)(DC)(DC)(DA)(DG) (DG)(DC)(DA)(DC) (DG)(DT)(DG)(DT)(DC)(DA)(DG)(DA)(DT)(DA) (DT)(DA)(DT)(DA)(DC)(DA) (DT)(DC)(DC) (DT)(DG)(DT)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.8 mg/mL
BufferpH: 7.5 / Details: 20mM Tris 7.5, 50mM NaCl, 1mM DTT, 1mM EDTA
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Average exposure time: 10.0 sec. / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 130000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: NONE
Final reconstructionNumber classes used: 1 / Resolution.type: BY AUTHOR / Resolution: 2.58 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 3.2.0) / Number images used: 302427
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.2.0)
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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Atomic model buiding 1

RefinementSpace: REAL / Protocol: AB INITIO MODEL
Output model

PDB-8cze:
Structure of a Xenopus Nucleosome with Widom 601 DNA

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